Analysis and/or measurement of an objective component to be measured by measuring luminescence, fluorescence, phosphorescence or the like have been widely utilized especially for the measurement of trace components such as environmental hormones or in vivo substances, because of its high sensitivity. In such highly sensitive spectrophotometry, small signal variation is required.
However, there is a problem of inability of correct measurement. Because a photodetector used for such high sensitive spectrophotometry is so sensitive, the photodetector detects even weak signals irrelevant to the objective measurement. As a method for preventing such measurement errors caused by weak signals irrelevant to the objective measurement, for example, a method for avoiding the influence of incorrect electric current by grounding of the photoelectric converter, or a method for detecting only objective signals by subtracting incorrect current measured in advance has been employed.
However, even though these procedures are performed, problems which are disturbed the high-accuracy measurement of the objective component remains, since background value varies by each measurement. Therefore, correct values cannot be obtained unless a calibration curve showing the relationship between the signal values and the concentrations of the objective component obtained by measuring the signal value using standard of the objective component containing known concentrations as a sample is established for each measurement.
For example, when a trace component is determined by using a luminescence property of luminol, luminol is oxidized by an action of appropriate amount of an oxidase such as peroxidase (POD) in accordance with the amount of the objective component to be measured in the presence of hydrogen peroxide, and fluorescence is generated (emit light). However, even when the concentration of the objective component is zero and the oxidase such as POD is not present, luminol is oxidized and generates luminescence slightly. There are problems which these energy variation value (background value) of luminol caused by factors other than the oxidase such as POD, or energy variation value generated by the oxidase such as POD (they are often referred to collectively as “signal value”), are varied significantly by change in measurement environment such as day and time of measurement, measurement place, and the like.
Such phenomena as mentioned above are generally observed in a method for measuring a signal value of energy variation of a substance such as luminescence, fluorescence, phosphorescence, and thus posing problems.
Therefore, development of a measuring method which provides low variation of the signal values and an instrument used for the method has been desired.